The aim of the proposed research is to analyze the structures and elucidate the functions of two major nucleolar nonribosomal, nonhistone proteins, B23 (MW 38,000, pI 5.1) and C23 (MW 103,000, pI 5.2). The two proteins are believed to function as organizer molecules for various components of the nucleolus. They represent a new class of nuclear protein, the "negative cluster proteins" which contain highly acidic, phosphorylated regions. The proteins are associated with nucleolar preribosomal particles and they bind silver under the same conditions that stain the nucleolus organizer regions of chromosomes. Protein C23 also binds DNA and a fraction of it is tightly bound to chromatin. The proteins will be isolated and subjected to automated and conventional sequence analyses. The major goal of the study is to obtain structural information on the two proteins, including: (1) the numbers and structures of the highly negatively charged clusters, (2) the distribution of the clusters along the polypeptide chain, (3) the sequences surrounding sites of the phosphorylation and (4) the eventual determination of the total sequences. To learn more about the functions of proteins C23 and B23 additional goals include: (1) localization and quantitation of the two proteins in various preribosomal particles and in subfractions of chromatin, (2) determination of the relative turnover rates to ascertain if they are "nucleolar stable" proteins and (3) studies of the DNA binding properties of the proteins to determine whether they recognize specific sites on DNA. The studies are intended as an early step in the understanding of the relationship of structure to function in the proteins involved in ribosome biogenesis.